A fine structure restriction endonuclease cleavage map of mitochondrial DNA (mtDNA) from mouse L cells has been derived using mtDNA sequences amplified in recombinant plasmids. L cell mutants resistant to chloramphenicol, oligomycin and antimycin-A will be tested for cytoplasmic inheritance of drug resistance and then screened for mtDNA sequence changes which result in alteration in cleavage patterns. mtDNA from the mutant cells will be cloned, amplified in E. coli and used to develop a transfomation system in which the purified mtDNA can be reintroduced into drug-sensitive L cells and expressed. Liposomes and erythrocyte ghosts will be used to package DNA in these experiments in an effort to avoid the passage through lysosomes which occurs in current mammalian cell transformation procedures. The mitochondrial ATPase of oligomycin-sensitive and resistant cells will be isolated and characterized in an attempt to understand the biochemical basis for drug resistance. Respiration defective mutants of Chinese hamster fibroblasts will also be examined for defects in gene products of mtDNA and mtDNA sequence alterations. Considerable intraspecies sequence heterogeneity has been found in mtDNA from natural populations of the field mouse Peromyscus. Similar sequence differences will be looked for in other mammals. We believe that mtDNA sequences can be used to follow gene flow within populations.